REASSESSING THE ROLE OF HOMOCYSTEINE AND HOLOTRANSCOBALAMIN LEVELS IN DIAGNOSING VITAMIN B12 DEFICIENCY ANEMIA

Background: Vitamin B12 deficiency can lead to irreversible neurological damage, megaloblastic anemia, osteoporosis, cerebrovascular, and cardiovascular diseases, and thus, early diagnosis is essential. Objective: The objective of the study was to assess homocysteine (Hcy) and holotranscobalamine (HoloTC) levels among patients with Vitamin B12 deficiency and to see if Hcy and HoloTC level assay can help us in diagnosis of Vitamin B12 deficiency. Methods: We carried out a cross-sectional observational study on 60 patients of Sr. B12 deficient male and female patients between the ages of 18 and 65 years in IPD and OPD patients at multispecialty hospital. Data were collected from predefined pro forma and were asked about their diet, socioeconomic status, and history. Then, these patients further undergone anthropometric measurements and investigated for Hcy and HoloTC level. The statistical analysis was done using Statistical Package for the Social Sciences (SSPS) software (version 11). Results: About 60% of cases were vegetarian and 40% of cases were non-vegetarian. Mean age of study participants was 43.67 years, mean of mean cellular volume was 90.7 fl, mean of B12 was 138 pmol/L, mean of HoloTC was 60.84 pmol/L, and mean of Hcy was 34.17 umol/L. Out of 60 patients, 10 patients had anemia, 21 patients had neurological manifestation, and 29 patients had gastrointestinal (GI) manifestation. In male group, out of 32, 11 patients had HoloTC <8.9, 19 patients had value between 8.9 and 128, and two patients had HoloTC more than 128. In female group, out of 28, seven patients had HoloTC <8.9, 14 patients had value between 8.9 and 128, and seven patients had HoloTC >128. In group of 32 male patients, none of male patients showed Hcy value <5.9, four patients showed Hcy between 5.9 and 16, and 28 patients showed Hcy value >16. Out of 28 female patients, none of female patients showed Hcy <3.36, nine patients showed Hcy between 3.36 and 20.4, and 19 patients showed Hcy >20.4. p <0.001 is highly statistically significant. Conclusion: In our study, we found that 31.33% of cases also showed decreased HoloTC along with B12 deficiency, but this correlation was statistically insignificant. We also found that 78.33% of cases showed increased Hcy along with serum B12 deficiency, which was statistically significant, so we concluded that there is a strong association between serum B12 and Hcy. We found that all patients with elevated Hcy also had low HoloTC except in two cases, but this correlation was not found to be statistically significant

“EFFECT OF BODY MASS INDEX ON PREGNANCY OUTCOME” - A PROSPECTIVE STUDY

Background: Mothers who are overweight or obese during pregnancy and childbirth, are known to be at risk of significant antenatal, intrapartum, postpartum, and neonatal complications. Objectives: The objective of the study was to evaluate the impact of high pre pregnancy body mass index (BMI) (<12 weeks of gestation) on the occurrence of maternal pregnancy outcome. A longitudinal observational study was carried out in a tertiary care hospital. In Group I, 50 antenatal women with gestational age <12 weeks BMI 18.5–35 kg/m2 and having singleton pregnancies were included in the study, while 50 women with normal BMI formed the Group II. Both groups were followed up throughout pregnancy and post-natal to assess complication during pregnancy, labor, and puerperium. Results: The mean BMI in Group I and Group II was 27.516 kg/m2 and 21.433 kg/m2. The prevalence of anemia was 40% and 26% among two groups. Antenatal and post-natal complications were gestational diabetes mellitus (Group I - 28% and Group II - 6%), preeclampsia (Group I - 16% and Group II - 2%), required induction of labor (Group I - 26% and Group II - 6%), preterm labor (Group I - 4% and Group II - 16%), and meconium staining of liquor (GroupI-20% and GroupII-12%), and the difference was statistically significant among two groups. Newborn complications were weight ≥2.5 kg (Group I - 74% and Group II - 48%), neonatal intensive care unit admission requirement (Group I - 26% and Group II - 17%), and the difference was statistically significant among two groups. Other complications which were not statistically significant among two groups were oligohydramnios (Group I - 2% and Group II - 4%), polyhydramnios (Group I - 6% and Group II - 4%), and appearance, pulse, grimace, activity, and respiration score at 1 min <7 (Group I - 14% and Group II - 6%). Conclusion: Pregnancy complications related to maternal BMI is a growing problem. Both lean and obese mothers carry an increased risk of adverse perinatal outcome. Given the major economic and medical consequence of pregnancy in these women, all attempts should be made to maintain a normal BMI in women of childbearing age. Pre-pregnancy counseling, health programs and appropriate multidisciplinary management should be done

Circulating Zonulin, a Marker of Intestinal Permeability, Is Increased in Association with Obesity-Associated Insulin Resistance

Zonulin is the only physiological mediator known to regulate intestinal permeability reversibly by modulating intercellular tight junctions. To investigate the relationship between intestinal permeability and obesity-associated metabolic disturbances in humans, we aimed to study circulating zonulin according to obesity and insulin resistance. Circulating zonulin (ELISA) was measured in 123 caucasian men in association with inflammatory and metabolic parameters (including minimal model-measured insulin sensitivity). Circulating zonulin increased with body mass index (BMI), waist to hip ratio (WHR), fasting insulin, fasting triglycerides, uric acid and IL-6, and negatively correlated with HDL-cholesterol and insulin sensitivity. In multiple regression analysis, insulin sensitivity (p = 0.002) contributed independently to circulating zonulin variance, after controlling for the effects of BMI, fasting triglycerides and age. When circulating IL-6 was added to this model, only BMI (p = 0.01) contributed independently to circulating zonulin variance. In conclusion, the relationship between insulin sensitivity and circulating zonulin might be mediated through the obesity-related circulating IL-6 increase

A study on trypsin, Aspergillus flavus and Bacillus sp. protease inhibitory activity in Cassia tora (L.) syn Senna tora (L.) Roxb. seed extract

<p>Abstract</p> <p>Background</p> <p>Proteases play an important role in virulence of many human, plant and insect pathogens. The proteinaceous protease inhibitors of plant origin have been reported widely from many plant species. The inhibitors may potentially be used for multiple therapeutic applications in viral, bacterial, fungal diseases and physiological disorders. In traditional Indian medicine system, <it>Cassia tora </it>(<it>Senna tora</it>) is reportedly effective in treatment of skin and gastrointestinal disorders. The present study explores the protease inhibitory activity of the above plant seeds against trypsin, <it>Aspergillus flavus </it>and <it>Bacillus </it>sp. proteases.</p> <p>Methods</p> <p>The crushed seeds of <it>Cassia tora </it>were washed thoroughly with acetone and hexane for depigmentation and defatting. The proteins were fractionated by ammonium sulphate (0-30, 30-60, 60-90%) followed by dialysis and size exclusion chromatography (SEC). The inhibitory potential of crude seed extract and most active dialyzed fraction against trypsin and proteases was established by spot test using unprocessed x-ray film and casein digestion methods, respectively. Electrophoretic analysis of most active fraction (30-60%) and SEC elutes were carried employing Sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) and Gelatin SDS-PAGE. Inhibition of fungal spore germination was studied in the presence of dialyzed active inhibitor fraction. Standard deviation (SD) and ANOVA were employed as statistical tools.</p> <p>Results</p> <p>The crude seeds' extract displayed strong antitryptic, bacterial and fungal protease inhibitory activity on x-ray film. The seed protein fraction 30-60% was found most active for trypsin inhibition in caseinolytic assay (P < 0.001). The inhibition of caseinolytic activity of the proteases increased with increasing ratio of seed extract. The residual activity of trypsin, <it>Aspergillus flavus </it>and <it>Bacillus </it>sp. proteases remained only 4, 7 and 3.1%, respectively when proteases were incubated with 3 mg ml^-1seed protein extract for 60 min. The inhibitory activity was evident in gelatin SDS-PAGE where a major band (~17-19 kD) of protease inhibitor (PI) was detected in dialyzed and SEC elute. The conidial germination of <it>Aspergillus flavus </it>was moderately inhibited (30%) by the dialyzed seed extract.</p> <p>Conclusions</p> <p><it>Cassia tora </it>seed extract has strong protease inhibitory activity against trypsin, <it>Aspergillus flavus </it>and <it>Bacillus </it>sp. proteases. The inhibitor in <it>Cassia tora </it>may attenuate microbial proteases and also might be used as phytoprotecting agent.</p

Networked T Cell Death following Macrophage Infection by Mycobacterium tuberculosis

<div><h3>Background</h3><p>Depletion of T cells following infection by <em>Mycobacterium tuberculosis</em> (Mtb) impairs disease resolution, and interferes with clinical test performance that relies on cell-mediated immunity. A number of mechanisms contribute to this T cell suppression, such as activation-induced death and trafficking of T cells out of the peripheral circulation and into the diseased lungs. The extent to which Mtb infection of human macrophages affects T cell viability however, is not well characterised.</p> <h3>Methodology/Principal Findings</h3><p>We found that lymphopenia (<1.5×10⁹ cells/l) was prevalent among culture-positive tuberculosis patients, and lymphocyte counts significantly improved post-therapy. We previously reported that Mtb-infected human macrophages resulted in death of infected and uninfected bystander macrophages. In the current study, we sought to examine the influence of infected human alveolar macrophages on T cells. We infected primary human alveolar macrophages (the primary host cell for Mtb) or PMA-differentiated THP-1 cells with Mtb H37Ra, then prepared cell-free supernatants. The supernatants of Mtb-infected macrophages caused dose-dependent, caspase-dependent, T cell apoptosis. This toxic effect of infected macrophage secreted factors did not require TNF-α or Fas. The supernatant cytotoxic signal(s) were heat-labile and greater than 50 kDa in molecular size. Although ESAT-6 was toxic to T cells, other Mtb-secreted factors tested did not influence T cell viability; nor did macrophage-free Mtb bacilli or broth from Mtb cultures. Furthermore, supernatants from <em>Mycobacterium bovis</em> Bacille de Calmette et Guerin (BCG)- infected macrophages also elicited T cell death suggesting that ESAT-6 itself, although cytotoxic, was not the principal mediator of T cell death in our system.</p> <h3>Conclusions</h3><p>Mtb-Infected macrophages secrete heat-labile factors that are toxic to T cells, and may contribute to the immunosuppression seen in tuberculosis as well as interfere with microbial eradication in the granuloma.</p> </div

Immunogenicity and Efficacy of Single Antigen Gp63, Polytope and PolytopeHSP70 DNA Vaccines against Visceral Leishmaniasis in Experimental Mouse Model

Polytope approach of genetic immunization is a promising strategy for the prevention of infectious disease as it is capable of generating effective cell mediated immunity by delivering the T cell epitopes assembled in series. Leishmaniasis is a significant world wide health problem for which no vaccine exists. In this study we have compared immunogenicity and efficacy of three types of DNA vaccines: single antigen Gp63 (Gp63/pcDNA), polytope (Poly/pcDNA) and Polytope fused with hsp70 (Poly/hsp/pcDNA) against visceral leishmaniasis in susceptible BALB/c mice. Mice vaccinated with these plasmids generated strong Th1 immune response as seen by dominating IFN-γ over IL-10 cytokine. Interestingly, cytotoxic responses generated by polytope DNA plasmid fused with hsp70 of Leishmania donovani were significantly higher when compared to polytope and single antigen Gp63 vaccine. Challenge studies revealed that the parasite load in liver and spleen was significantly lower with Poly/hsp/pcDNA vaccination compared to other vaccines. Therefore, our study indicates that polytope DNA vaccine is a feasible, practical and effective approach for visceral leishmaniasis

Comparative Functional Genomics of Salt Stress in Related Model and Cultivated Plants Identifies and Overcomes Limitations to Translational Genomics

One of the objectives of plant translational genomics is to use knowledge and genes discovered in model species to improve crops. However, the value of translational genomics to plant breeding, especially for complex traits like abiotic stress tolerance, remains uncertain. Using comparative genomics (ionomics, transcriptomics and metabolomics) we analyzed the responses to salinity of three model and three cultivated species of the legume genus Lotus. At physiological and ionomic levels, models responded to salinity in a similar way to crop species, and changes in the concentration of shoot Cl− correlated well with tolerance. Metabolic changes were partially conserved, but divergence was observed amongst the genotypes. Transcriptome analysis showed that about 60% of expressed genes were responsive to salt treatment in one or more species, but less than 1% was responsive in all. Therefore, genotype-specific transcriptional and metabolic changes overshadowed conserved responses to salinity and represent an impediment to simple translational genomics. However, ‘triangulation’ from multiple genotypes enabled the identification of conserved and tolerant-specific responses that may provide durable tolerance across species

Leishmania donovani: Immunostimulatory Cellular Responses of Membrane and Soluble Protein Fractions of Splenic Amastigotes in Cured Patient and Hamsters

Visceral leishmaniasis (VL), caused by the intracellular parasite Leishmania donovani, L. chagasi and L. infantum is characterized by defective cell-mediated immunity (CMI) and is usually fatal if not treated properly. An estimated 350 million people worldwide are at risk of acquiring infection with Leishmania parasites with approximately 500,000 cases of VL being reported each year. In the absence of an efficient and cost-effective antileishmanial drug, development of an appropriate long-lasting vaccine against VL is the need of the day. In VL, the development of a CMI, capable of mounting Th1-type of immune responses, play an important role as it correlate with recovery from and resistance to disease. Resolution of infection results in lifelong immunity against the disease which indicates towards the feasibility of a vaccine against the disease. Most of the vaccination studies in Leishmaniasis have been focused on promastigote- an infective stage of parasite with less exploration of pathogenic amastigote form, due to the cumbersome process of its purified isolation. In the present study, we have isolated and purified splenic amastigotes of L. donovani, following the traditional protocol with slight modification. These were fractionated into five membranous and soluble subfractions each i.e MAF1-5 and SAF1-5 and were subjected for evaluation of their ability to induce cellular responses. Out of five sub-fractions from each of membrane and soluble, only four viz. MAF2, MAF3, SAF2 and SAF3 were observed to stimulate remarkable lymphoproliferative, IFN-γ, IL-12 responses and Nitric Oxide production, in Leishmania-infected cured/exposed patients and hamsters. Results suggest the presence of Th-1 type immunostimulatory molecules in these sub-fractions which may further be exploited for developing a successful subunit vaccine from the less explored pathogenic stage against VL

Silencing of Vlaro2 for chorismate synthase revealed that the phytopathogen Verticillium longisporum induces the cross-pathway control in the xylem

The first leaky auxotrophic mutant for aromatic amino acids of the near-diploid fungal plant pathogen Verticillium longisporum (VL) has been generated. VL enters its host Brassica napus through the roots and colonizes the xylem vessels. The xylem contains little nutrients including low concentrations of amino acids. We isolated the gene Vlaro2 encoding chorismate synthase by complementation of the corresponding yeast mutant strain. Chorismate synthase produces the first branch point intermediate of aromatic amino acid biosynthesis. A novel RNA-mediated gene silencing method reduced gene expression of both isogenes by 80% and resulted in a bradytrophic mutant, which is a leaky auxotroph due to impaired expression of chorismate synthase. In contrast to the wild type, silencing resulted in increased expression of the cross-pathway regulatory gene VlcpcA (similar to cpcA/GCN4) during saprotrophic life. The mutant fungus is still able to infect the host plant B. napus and the model Arabidopsis thaliana with reduced efficiency. VlcpcA expression is increased in planta in the mutant and the wild-type fungus. We assume that xylem colonization requires induction of the cross-pathway control, presumably because the fungus has to overcome imbalanced amino acid supply in the xylem